Integrin Alpha v Beta 6 (αvß6) and Its Implications in Cancer Treatment.

Integrins are necessary for cell adhesion, migration, and positioning. Essential for inducing signalling events for cell survival, proliferation, and differentiation, they also trigger a variety of signal transduction pathways involved in mediating invasion, metastasis, and squamous-cell carcinoma. Several recent studies have demonstrated that the up- and down-regulation of the expression of αv and other integrins can be a potent marker of malignant diseases and patient prognosis. This review focuses on an arginine-glycine-aspartic acid (RGD)-dependent integrin αVß6, its biology, and its role in healthy humans. We examine the implications of αVß6 in cancer progression and the promotion of epithelial-mesenchymal transition (EMT) by contributing to the activation of transforming growth factor beta TGF-ß. Although αvß6 is crucial for proper function in healthy people, it has also been validated as a target for cancer treatment. This review briefly considers aspects of targeting αVß6 in the clinic via different therapeutic modalities.

New Insights on the Feature and Function of Tail Tubular Protein B and Tail Fiber Protein of the Lytic Bacteriophage φYeO3-12 Specific for Yersinia enterocolitica Serotype O:3.

For the first time, we are introducing TTPBgp12 and TFPgp17 as new members of the tail tubular proteins B (TTPB) and tail fiber proteins (TFP) family, respectively. These proteins originate from Yersinia enterocolitica phage φYeO3-12. It was originally thought that these were structural proteins. However, our results show that they also inhibit bacterial growth and biofilm formation. According to the bioinformatic analysis, TTPBgp12 is functionally and structurally similar to the TTP of Enterobacteria phage T7 and adopts a ß-structure. TFPgp17 contains an intramolecular chaperone domain at its C-terminal end. The N-terminus of TFPgp17 is similar to other representatives of the TFP family. Interestingly, the predicted 3D structure of TFPgp17 is similar to other bacterial S-layer proteins. Based on the thermal unfolding experiment, TTPBgp12 seems to be a two-domain protein that aggregates in the presence of sugars such as maltose and N-acetylglucosamine (GlcNAc). These sugars cause two unfolding events to transition into one global event. TFPgp17 is a one-domain protein. Maltose and GlcNAc decrease the aggregation temperature of TFPgp17, while the presence of N-acetylgalactosamine (GalNAc) increases the temperature of its aggregation. The thermal unfolding analysis of the concentration gradient of TTPBgp12 and TFPgp17 indicates that with decreasing concentrations, both proteins increase in stability. However, a decrease in the protein concentration also causes an increase in its aggregation, for both TTPBgp12 and TFPgp17.

Effectiveness of Sensors Contact Metallization (Ti, Au, and Ru) and Biofunctionalization for Escherichia coli Detection.

In designing a bacteria biosensor, various issues must be addressed: the specificity of bacteria recognition, the immobilization of biomolecules that act as the bacteria receptor, and the selectivity of sensor surface. The aim of this paper was to examine how the biofunctionalized surface of Ti, Au, and Ru metals reacts in contact with strains of Escherichia coli (E. coli). The focus on metal surfaces results from their future use as electrodes in high frequency biosensors, e.g., resonant circuits or transmission-line sections. First, the surfaces of different metals were chemically functionalized with 3-aminopropyltriethoxysilane (APTES) and glutaraldehyde or with 3-glycidylooxypropyltrimethoxysilane (GPTMS) followed by N-(5-amino-1-carboxypentyl) iminodiacetic acid (AB-NTA) and NiCl2. Secondly, the lipopolysaccharide binding protein (LBP), polyclonal anti-Escherichia coli antibody and bacteriophage protein gp37 were tested as bacteria receptors. The selectivity and specificity have been confirmed by the Enzyme-Linked Immunosorbent Assay (ELISA) and visualized by scanning electron microscopy at low landing energies. We noticed that LBP, polyclonal antibody, and gp37 were successfully immobilized on all studied metals and recognized the E. coli bacteria selectively. However, for the antibody, the highest reactivity was observed when Ti surface was modified, whereas the bacteria binding was comparable between LBP and gp37 on the functionalized Ru surfaces, independent from modification. Thus, all surfaces were biocompatible within the scope of biosensor functionality, with titanium functionalization showing the best performance.

Pathogenic factors of Pseudomonas aeruginosa - the role of biofilm in pathogenicity and as a target for phage therapy.

Pseudomonas aeruginosa is an opportunistic pathogen that can cause several acute and chronic infections in humans, and it has become an important cause of nosocomial infections and antibiotic resistance. Biofilm represents an important virulence factor for these bacteria, plays a role in P. aeruginosa infections and avoidance of immune defence mechanisms, and has the ability to protect the bacteria from antibiotics. Alginate, Psl and Pel, three exopolysaccharides, are the main components in biofilm matrix, with many biological functions attributed to them, especially with respect to the protection of the bacterial cell from antibiotics and the immune system. Pseudomonas infections, biofilm formation and development of resistance to antibiotics all require better understanding to achieve the best results using alternative treatment with phage therapy. This review describes the P. aeruginosa pathogenicity and virulence factors with a special focus on the biofilm and its role in infection and resistance to antibiotics and summarizes phage therapy as an alternative approach in treatment of P. aeruginosa infections.

Chemical characterization and immunomodulatory properties of polysaccharides isolated from probiotic Lactobacillus casei LOCK 0919.

The Lactobacillus casei strain, LOCK 0919, is intended for the dietary management of food allergies and atopic dermatitis (LATOPIC® BIOMED). The use of a probiotic to modulate immune responses is an interesting strategy for solving imbalance problems of gut microflora that may lead to various disorders. However, the exact bacterial signaling mechanisms underlying such modulations are still far from being understood. Here, we investigated variations in the chemical compositions and immunomodulatory properties of the polysaccharides (PS), L919/A and L919/B, which are produced by L. casei LOCK 0919. By virtue of their chemical features, such PS can modulate the immune responses to third-party antigens. Our results revealed that L919/A and L919/B could both modulate the immune response to Lactobacillus planatarum WCFS1, but only L919/B could alter the response of THP-1 cells (in terms of tumor necrosis factor alpha production) to L. planatarum WCFS1 and Escherichia coli Nissle 1917. The comprehensive immunochemical characterization is crucial for the understanding of the biological function as well as of the bacteria-host and bacteria-bacteria cross-talk.

Identification of Lactobacillus proteins with different recognition patterns between immune rabbit sera and nonimmune mice or human sera.

BACKGROUND: The genus Lactobacillus belongs to a large heterogeneous group of low G + C Gram-positive anaerobic bacteria, which are frequently used as probiotics. The health-beneficial effects, in particular the immunomodulation effect, of probiotics depend on the strain and dose used. Strain variations may be related to diversity of the cell surface architecture of bacteria and the ability to express specific antigens or secrete compounds. The use of Lactobacillus as probiotic requires a comprehensive understanding of its effect on host immune system. To evaluate the potential immunoreactive properties of proteins isolated from four Lactobacillus strains: L. johnsonii 142 and L. johnsonii 151, L. rhamnosus LOCK 0900 and L. casei LOCK 0919, the polyclonal sera obtained from mouse and human have been tested as well as with sera from rabbits immunized with whole lactobacilli cells. RESULTS: The reactivity of isolated proteins detected by SDS-PAGE and Western blotting was heterogeneous and varied between different serum samples. The proteins with the highest immunoreactivity were isolated, purified and sequenced, in particular the fractions were identified as phosphoglycerate kinase (L. johnsonii 142), glyceraldehyde 3-phosphate dehydrogenase (L. johnosnii 142, L. rhamnosus LOCK 0900), hypothetic protein JDM1_1307 (L. johnsonii 151) and fructose/tagatose-bisphosphate-aldolase (L. casei LOCK 0919). CONCLUSION: The different prevalence of reactions against tested antigens in rabbit, mouse and human sera may indicate significant differences in immune system and commensal cross-talk in these groups. The identification of immunoreactive lactobacilli proteins opens the possibility to use them as an antigens for development of vaccines.

Label-free sensitivity of long-period gratings enhanced by atomic layer deposited TiO(2) nano-overlays.

In this paper, we discuss an impact of thin titanium dioxide (TiO(2)) coatings on refractive index (RI) sensitivity and biofunctionalization of long-period gratings (LPGs). The TiO(2) overlays on the LPG surfaces have been obtained using atomic layer deposition (ALD) method. This method allows for a deposition of conformal, thickness-controlled, with well-defined optical properties, and high-RI thin films which are highly desired for optical fiber sensors. It has been found that for LPGs working at a dispersion turning point of higher order cladding modes only tens of nanometers of TiO(2) overlay thickness allow to obtain cladding mode transition effect, and thus significant improvement of RI sensitivity. When the TiO(2) overlay thickness reaches 70 nm, it is possible to obtain RI sensitivity exceeding 6200 nm/RIU in RI range where label-free sensors operate. Moreover, LPGs with TiO(2)-enhanced RI sensitivity have shown improved sensitivity to bacteria endotoxin (E. coli B lipopolysaccharide) detection, when TiO(2) surface is functionalized with endotoxin binding protein (adhesin) of T4 bacteriophage.

Distinct immunomodulation of bone marrow-derived dendritic cell responses to Lactobacillus plantarum WCFS1 by two different polysaccharides isolated from Lactobacillus rhamnosus LOCK 0900.

The structures of polysaccharides (PS) isolated from Lactobacillus rhamnosus LOCK 0900 and results from stimulation of mouse bone marrow-derived dendritic cells (BM-DC) and human embryonal kidney (HEK293) cells stably transfected with Toll-like receptors (TLR) upon exposure to these antigens were studied. L. rhamnosus LOCK 0900 produces PS that differ greatly in their structure. The polymer L900/2, with a high average molecular mass of 830 kDa, is a branched heteropolysaccharide with a unique repeating unit consisting of seven sugar residues and pyruvic acid, whereas L900/3 has a low average molecular mass of 18 kDa and contains a pentasaccharide repeating unit and phosphorus. Furthermore, we found that both described PS neither induce cytokine production and maturation of mouse BM-DC nor induce signaling through TLR2/TLR4 receptors. However, they differ profoundly in their abilities to modulate the BM-DC immune response to the well-characterized human isolate Lactobacillus plantarum WCFS1. Exposure to L900/2 enhanced interleukin-10 (IL-10) production induced by L. plantarum WCFS1, while in contrast, L900/3 enhanced the production of IL-12p70. We conclude that PS, probably due to their chemical features, are able to modulate the immune responses to third-party antigens. The ability to induce regulatory IL-10 by L900/2 opens up the possibility to use this PS in therapy of inflammatory conditions, such as inflammatory bowel disease, whereas L900/3 might be useful in reverting the antigen-dependent Th2-skewed immune responses in allergies.

Application of non-parametric correlations to compare the compliance of Beighton and Sachse tests in the assessment of hypermobility based on research of the fitness instructors group.

BACKGROUND: Joint hypermobility is a spectrum of symptoms associated with connective tissue disorders. The main feature is the increased range of joint mobility. Hypermobility is rarely recognised in clinical practice. The diagnosis is based on the evaluation of diagnostic tests, mainly the Beighton score. The divergence of research methods means that patients do not receive a proper diagnosis and treatment. METHODS: The study used the Beighton score and the Sachse scale. Both tests are "all-or-none-tests". Non-parametric correlations were used to assess the concordance effect. To this end, two methods were adopted, i.e., the Spearman Rank Correlation and Kendall tau Rank Correlation. The values of correlation coefficients were calculated, respectively, rho and Kendall tau. The study involved 30 women working as fitness instructors. RESULTS: Consent results of hypermobility assessment for both methods were obtained in 3 cases, while the discrepancy in the hypermobility statement concerns measurements made in 10 participants. This cursory assessment already indicates a significant differentiation of results obtained for both methods. DISCUSSION: To the best of our knowledge, there are not many studies comparing different HSD diagnostic methods. The Beighton score is the most commonly used, but the selection of only 5 joints for the examination does not show the systemic nature of hypermobility. A reliable methodology should be based not only on goniometric measurements of selected joints. CONCLUSION: The expanded correlation analysis of Beighton and Sachse hypermobility tests indicates their poor compliance. Therefore, there is a need to standardise hypermobility spectrum disorder diagnostics, which may affect the objectification and credibility of these diagnostics.

[Biological properties of Lactobacillus surface proteins]. / Wlasciwosci biologiczne bialek powierzchniowych bakterii z rodzaju Lactobacillus.

Lactobacillus, a genus of Gram-positive bacteria, includes many strains of probiotic microflora. Probiotics, by definition, are living microorganisms that exert beneficial effects on the host organism. The morphology and physiology of the Lactobacillus bacterial genus are described. The structure of the cell wall of Gram-positive bacteria is discussed. The surface S-layer of Lactobacillus composed of proteins (SLP) with low molecular mass is presented. Cell surface proteins participating in the regulation of growth and survival of the intestinal epithelium cells are characterized. The influence of stress factors such as increased temperature, pH, and enzymes of gastric and pancreatic juice on SLP expression is described. The ability of binding of heavy metal ions by S-layer proteins is discussed. The characteristics of these structures, including the ability to adhere to epithelial cells, and the inhibition of invasion of pathogenic microflora of type Shigella, Salmonella, Escherichia coli and Clostridium and their toxins, are presented. 

[Surface proteins of bacteria of the genus Bifidobacterium]. / Bialka powierzchniowe bakterii z rodzaju Bifidobacterium.

Beneficial effects due to the presence of probiotic bacteria of the genus Bifidobacterium in the human intestinal tract are still an interesting object of study. So far activities have been confirmed of bifidobacteria in stimulation of the host immune system, stimulation of tumor cell apoptosis, improvement of bowel motility, alleviation of symptoms of lactose intolerance, cholesterol lowering capacity, prevention and treatment of diarrhea and irritable bowel syndrome, alleviation of allergy or atopic dermatitis, maintenance of homeostasis of the intestine, and stimulation of the development of normal intestinal microflora in infants. A multitude of therapeutic properties encourages researchers to investigate the possibility of using the potential of Bifidobacterium in the prevention and treatment of other conditions such as rheumatoid arthritis and depression. Although it is known that the beneficial effects are due to intestinal mucosal colonization by these bacteria, the cell components responsible for the colonization are still not determined. In addition to the beneficial effects of probiotic administration, there were also negative effects including sepsis. Therefore research has been directed to identify specific components of Bifidobacterium responsible for probiotic effects. Currently researchers are focused on identifying, isolating and evaluating the properties of surface proteins that are probably involved in the adhesion of bacterial cells to the intestinal epithelium, improving colonization. This paper is an overview of current knowledge on Bifidobacterium surface proteins. The ways of transport and anchoring proteins in Gram-positive bacterial cells, the assembly of cell wall, and a description of the genus Bifidobacterium are presented.

Detection and Level Evaluation of Antibodies Specific to Environmental Bacteriophage I11mO19 and Related Coliphages in Non-Immunized Human Sera.

Bacteriophages (phages) are viruses infecting bacteria. They are widely present in the environment, food, and normal microflora. The human microbiome is a mutually interdependent network of bacteria, bacteriophages, and human cells. The stability of these tri-kingdom interactions may be essential for maintaining immunologic and metabolic health. Phages, as with each other's antigens, may evoke an immune response during a human's lifetime and induce specific antibody generation. In this manuscript, we labeled these antibodies as naturally generated. Naturally generated antibodies may be one of the most important factors limiting the efficacy of phage therapy. Herein, we attempted to determine the physiological level of these antibodies specific to a population bacteriophage named I11mO19 in human sera, using an ELISA-based assay. First, we purified the phage particles and assessed the immunoreactivity of phage proteins. Then, affinity chromatography was performed on columns with immobilized phage proteins to obtain a fraction of human polyclonal anti-phage antibodies. These antibodies were used as a reference to elaborate an immunoenzymatic test that was used to determine the level of natural anti-phage antibodies. We estimated the average level of anti-I11mO19 phage antibodies at 190 µg per one milliliter of human serum. However, immunoblotting revealed that cross-reactivity occurs between some proteins of I11mO19 and two other coliphages: T4 and ΦK1E. The antigens probably share common epitopes, suggesting that the determined level of anti-I11mO19 phage might be overestimated and reflects a group of antibodies reactive to a broad range of other E. coli phages. Anti-I11mO19 antibodies did not react with Pseudomonas bacteriophage F8, confirming specificity to the coliphage group. In this work, we wanted to show whether it is possible to determine the presence and level of anti-phage antibodies in nontargeted-immunized sera, using an immunoenzymatic assay. The conclusion is that it is possible, and specific antibodies can be determined. However, the specificity refers to a broader coliphage group of phages, not only the single phage strain.

Bacteriophage Tail Proteins as a Tool for Bacterial Pathogen Recognition-A Literature Review.

In recent years, a number of bacterial detection methods have been developed to replace time-consuming culture methods. One interesting approach is to mobilize the ability of phage tail proteins to recognize and bind to bacterial hosts. In this paper, the authors provide an overview of the current methodologies in which phage proteins play major roles in detecting pathogenic bacteria. Authors focus on proteins capable of recognizing highly pathogenic strains, such as Acinetobacter baumannii, Campylobacter spp., Yersinia pestis, Pseudomonas aeruginosa, Listeria monocytogenes, Staphylococcus aureus, Enterococcus spp., Salmonella spp., and Shigella. These pathogens may be diagnosed by capture-based detection methods involving the use of phage protein-coated nanoparticles, ELISA (enzyme-linked immunosorbent assay)-based methods, or biosensors. The reviewed studies show that phage proteins are becoming an important diagnostic tool due to the discovery of new phages and the increasing knowledge of understanding the specificity and functions of phage tail proteins.

φYeO3-12 phage tail fiber Gp17 as a promising high specific tool for recognition of Yersinia enterocolitica pathogenic serotype O:3.

Yersiniosis is an infectious zoonotic disease caused by two enteropathogenic species of Gram-negative genus Yersinia: Yersinia enterocolitica and Yersinia pseudotuberculosis. Pigs and other wild and domestic animals are reservoirs for these bacteria. Infection is usually spread to humans by ingestion of contaminated food. Yersiniosis is considered a rare disease, but recent studies indicate that it is overlooked in the diagnostic process therefore the infections with this bacterium are not often identified. Reliable diagnosis of Yersiniosis by culturing is difficult due to the slow growth of the bacteria easily overgrown by other more rapidly growing microbes unless selec-tive growth media is used. Phage adhesins recognizing bacteria in a specific manner can be an excellent diagnostic tool, es-pecially in the diagnosis of pathogens difficult for culturing. In this study, it was shown that Gp17, the tail fiber protein (TFP) of phage φYeO3-12, specifically recognizes only the pathogenic Yersinia enterocolitica serotype O:3 (YeO:3) bacteria. The ELISA test used in this work confirmed the specific interaction of this protein with YeO:3 and demonstrated a promising tool for developing the pathogen recognition method based on phage adhesins.

Molecular Characteristic, Antibiotic Resistance, and Detection of Highly Immunoreactive Proteins of Group B Streptococcus Strains Isolated From Urinary Tract Infections in Polish Adults.

Group B streptococcus (GBS) is one of the uropathogens that causes urinary tract infections (UTIs). The aims of this article were molecular characterization, an analysis of antimicrobial susceptibility profiles, adherence to bladder endothelial cells, and the detection of immunoreactive proteins of 94 clinical strains of GBS isolated from adult Polish patients with UTI. Antibiotic susceptibilities were determined by disk diffusion. Serotyping and Alp family genes detection were studied using multiplex PCR. Genetic profiles were determined by pulsed-field gel electrophoresis. The adherence ability of the studied strains was estimated by incubation on human bladder microvascular endothelial cell line. Immunoreactive proteins were studied by immunoblotting. Antibiotic susceptibility investigation revealed that 22% of GBS strains were resistant to erythromycin, whereas 18% demonstrated resistance to clindamycin. cMLSB was present in 76% of the resistant strains, M phenotype was detected in 14%, whereas iMLSB was present for 10%. The most common serotype was serotype III (31%), followed by serotype V (27%), and serotype Ia (17%). The genes that dominated among other Alp genes were: epsilon (29%), alp2 (27%), and rib (23%). The most common co-occurring serotypes and Alp genes were: Ia and epsilon, III and rib, III and alp2, V and alp2, and V and alp3 (p < 0.001). The PFGE method showed high clonality for serotype V and cMLSB (p < 001). The PFGE method showed high clonality for serotype V. Furthermore, this serotype was significantly associated with the cMLSB phenotype (p < 0.001). The most common immunoreactive proteins demonstrated masses of 50 kDa and 45-47 kDa. Although examined GBS isolates showed high genetic diversity, immunoreactive proteins were common for most of the studied GBS isolates, which may indicate their conservation, and allows to consider them as potential immunodiagnostic markers. Although the examined GBS isolates showed high genetic diversity, immunoreactive proteins were shared by most of the studied GBS isolates. It may indicate their conservation, thus allowing to consider them as potential immunodiagnostic markers.

[The functions of bacteriophage proteins]. / Funkcje bialek bakteriofagowych.

Bacteriophages (phages) are viruses specific towards bacterial strains and are natural regulators of bacterial populations in nature. Interest in the therapeutic use of phages is growing due to the emergence of antibiotic resistant pathogens. Bacteriophage proteins are responsible for phage specificity and virulence. These proteins are i) adhesins, capable of recognizing specific receptors on the bacterial surface, ii) enzymes responsible for destroying bacterial cell wall components or bacterial slime, and iii) structural proteins making up the phage capsid. Bacteriophage enzymes destroy the bacterial cell wall from both outside and inside by hydrolyzing carbohydrate and protein components. All these proteins protect phage genetic material, secure injection of the phage nucleic acid into the bacterial cell, and promote phage propagation. Our better understanding of how bacterial viruses function may help to combat antibiotic resistant human pathogens and expand the use of bacteriophages to animal and plant care. Other applications may include food processing and environmental protection.

Application of selected biosensor techniques in clinical diagnostics.

INTRODUCTION: Examination of disease biomarkers mostly performed on crude materials, such as serum, meets some obstacles, resulting from sample complexity and the wide range of concentrations and sizes of the components. Techniques currently used in clinical diagnostics are usually time-consuming and expensive. The more sensitive and portable devices are needed for early diagnostics. Chemical sensors are devices that convert chemical information into parameters suitable for fast and precise processing and measurement. AREA COVERED: We review the use of biosensors and their possible application in early diagnostics of some diseases like cancer or viral infections. We focus on different types of biorecognition and some technical modifications, lowering the limit of detection potentially attractive to medical practitioners. EXPERT OPINION: Among the new diagnostic strategies, the use of biosensors is of increasing interest. In these techniques, the capture ligand interacts with the analyte of interest. Measuring interactions between partners in real time by surface plasmon resonance yields valuable information about kinetics and affinity in a short time and without labels. Importantly, the tendency in such techniques is to make biosensor devices smaller and the test results apparent with the naked eye, so they can be used in point-of-care medicine.

Biochemical features of the novel Tail Tubular Protein A of Yersinia phage phiYeO3-12.

Tail Tubular Protein A (TTPA) was long thought to be strictly a structural protein of environmental bacteriophages. However, our recent work has suggested that some TTPAs have additional functional features and thus are dual-function proteins. This study introduces a new TTPA family member, TTPAgp11, which belongs to Yersinia phage phiYeO3-12. We cloned the gene, expressed it and then purified the phage protein. The protein, including its hydrolytic activity, was characterized. Our enzymatic activity tests showed that TTPAgp11 displayed hydrolytic activity towards Red-starch, suggesting that this enzyme could be classified as part as the α - 1, 4-glucosidase family. Protein folding and aggregation tests indicated that TTPAgp11 is a single-domain protein whose aggregation can be induced by maltose or N-acetylglucosamine. The spatial structure of TTPAgp11 seemed to resemble that of the first reported dual-function TTPA, TTPAgp31, which was isolated from Klebsiella pneumoniae phage 32.

The Podovirus ϕ80-18 Targets the Pathogenic American Biotype 1B Strains of Yersinia enterocolitica.

We report here the complete genome sequence and characterization of Yersinia bacteriophage vB_YenP_ϕ80-18. ϕ80-18 was isolated in 1991 using a Y. enterocolitica serotype O:8 strain 8081 as a host from a sewage sample in Turku, Finland, and based on its morphological and genomic features is classified as a podovirus. The genome is 42 kb in size and has 325 bp direct terminal repeats characteristic for podoviruses. The genome contains 57 predicted genes, all encoded in the forward strand, of which 29 showed no similarity to any known genes. Phage particle proteome analysis identified altogether 24 phage particle-associated proteins (PPAPs) including those identified as structural proteins such as major capsid, scaffolding and tail component proteins. In addition, also the DNA helicase, DNA ligase, DNA polymerase, 5'-exonuclease, and the lytic glycosylase proteins were identified as PPAPs, suggesting that they might be injected together with the phage genome into the host cell to facilitate the take-over of the host metabolism. The phage-encoded RNA-polymerase and DNA-primase were not among the PPAPs. Promoter search predicted the presence of four phage and eleven host RNA polymerase -specific promoters in the genome, suggesting that early transcription of the phage is host RNA-polymerase dependent and that the phage RNA polymerase takes over later. The phage tolerates pH values between 2 and 12, and is stable at 50°C but is inactivated at 60°C. It grows slowly with a 50 min latent period and has apparently a low burst size. Electron microscopy revealed that the phage has a head diameter of about 60 nm, and a short tail of 20 nm. Whole-genome phylogenetic analysis confirmed that ϕ80-18 belongs to the Autographivirinae subfamily of the Podoviridae family, that it is 93.2% identical to Yersinia phage fHe-Yen3-01. Host range analysis showed that ϕ80-18 can infect in addition to Y. enterocolitica serotype O:8 strains also strains of serotypes O:4, O:4,32, O:20 and O:21, the latter ones representing similar to Y. enterocolitica serotype O:8, the American pathogenic biotype 1B strains. In conclusion, the phage ϕ80-18 is a promising candidate for the biocontrol of the American biotype 1B Y. enterocolitica.

In silico analysis of bacteriophage tail tubular proteins suggests a putative sugar binding site and a catalytic mechanism.

Bacteriophage base tailplate proteins were recently discovered to have hydrolytic activity towards disaccharides. The putative assignment of sugar binding sites was based on known lectin structures and identified residues a.a. 40-120 as the potential binding region for disaccharides [1]. To help verify the prediction, an in silico analysis was performed on the structure of a base tailplate protein gp31 from Klebsiella pneumoniae bacteriophage KP32 (PDB: 5MU4) which shows activity towards maltose but not trehalose [1]. Based on the information, a full surface docking was performed for both sugars which identified 2 regions different than originally predicted. The first region clearly favored maltose during the docking phase while the second one allowed for the energetically-equivalent binding of trehalose. To verify the assignment, a molecular dynamics simulation was performed to assess the stability of the docked substrates. MD simulations suggested that the first site included residues D131, D133, and E134, and was also superior for maltose binding while clearly disfavoring trehalose. Analysis of the putative catalytic mechanism suggested residues D131, D133 and E134 as critical for substrate binding. The residue D133 did participate in a stable substrate binding and was positioned near the scissile bond, potentially making it a catalytic residue. Catalytic residues were most likely D131 and D133, one of the two options proposed by Pyra et al. [1]. A comparison with known hydrolase mechanisms suggested that the enzyme most likely retains configuration during hydrolysis of maltose. The findings are discussed for other bacteriophage proteins regarding their potential specificities and catalytic mechanisms.